The invention relates to novel neuropeptides, more specifically scorpion toxin-related peptides which are obtained from the venom gland of Heterometrus spinnifer, a species of scorpion.
xe2x80x9cScorpionxe2x80x9d is a general term for arthropods belonging to the class Arachnida, the order Scorpionida and about 600 species of scorpions now inhabit the world. Scorpions have generally believed to be virulently poisonous; however, actually those having fetal toxins are limited to only several species, including Buthus ustralis (living in the deserts of Africa), Centruroides exilicauda (native to Mexico) and B. occitauda (native to South Europe), etc. Up to now, various types of scorpion toxins have been isolated (Dreyer. F., Rev. Physiol. Biochem. Pharmacol., vol.15, pp.94-128, 1990), which are considered to be neurotoxins.
From the studies on pharmacological properties of these toxins, the actions of the toxins against K+ channels have been drawing attention. For example, margatoxin (MgTX) isolated from the venom gland of Centruroides margaritatus (Garcia-Calvo et al., J. Biol. Chem., vol. 268, pp. 18866-18874, 1993) and agitoxin 2 (AgTX2) isolated from the venom gland of Leiurus quinquestriatus herbraes (Garcia et al., Biochemistry, vol. 33, pp. 6834-6839, 1994) have been known to act as blockers on voltage-gated K+ channels. These toxins are peptides composed of 38 amino acid residues and containing three disulfide bridges in their molecules. Recently, also reported is a peptide, Pandinus imperator toxin 1 (Pi1), which is a peptide containing four disulfide bridges in the molecule and composed of 35 amino acid residues (Olamendi-Portugal et al., Biochem. J. vol. 315, pp. 977-981, 1996).
On the other hand, it has been considered that production of interleukin-2 (IL-2) by T cells requires Ca2+ influx into the T cells. Recently, it has been discovered that Kv1.3, which is one type of the K+ channels, is involved in Ca2+ influx into T cells accompanied by activation of the T cells (Leonard et al., Proc. Natl. Acad. Sci. USA, vol. 89, pp. 10094-10098, 1992). Further, Lin et al. reported that inhibition of Kv1.3 channels suppresses Ca2+ influx into T cells, proliferation of T cells and production of IL-2 by T cells (J. Exp. Med., vol. 177, pp.637-645, 1993).
Under these circumstances, scorpion toxins are expected to be applicable to medicines in view of their various pharmacological activities. However, for application to medicines, it is necessary to separate the useful pharmacological activities of the toxins from their undesirable toxicity and, consequently, it becomes necessary to isolate much scorpion toxins and clarify the structure-activity relationship thereof. In these situations, the object of the present invention is to provide novel scorpion toxin-related peptides for contribution to application of scorpion toxins to medicines.
The inventors have made intensive and extensive studies on isolation of novel scorpion toxin-related peptides from the venom gland of a scorpion Heterometrus spinnifer, on the basis of the ability to block the rat brain voltage-gated K+ channels Kv1.3 (also called xe2x80x9cRCK3xe2x80x9d) expressed in Xenopus oocytes. As a result, the inventors have succeeded in isolation and purification of a novel peptide, named HsTX1, represented by SEQ ID NO:1 in which four disulfide bridges are present in the molecule. They have also succeeded in determination of the primary and higher-order structures of HsTX1. It has been found that HsTX1 exhibits much stronger ability to block the K+ channels compared with the previously reported scorpion toxins and inhibits the IL-2 production by human peripheral blood T cells. Thus, the invention has been accomplished.
According to the present invention, there is provided a peptide represented by an amino acid sequence:
in which 0 to 4 disulfide bridges are present in the molecule and the C terminal may be amidated, as a blocker of voltage-gated K+ channels or an inhibitor of IL-2 production.
In a specific embodiment, the present invention provides a peptide represented by the above amino acid sequence in which the disulfide bridges are selected from the group consisting of CyS3-Cys24, Cys9-Cys29, Cys13, Cys31 and Cys19-Cys34.
In a most preferred embodiment, the present invention provides a peptide represented by the above amino acid sequence in which the C terminal is amidated and four disulfide bridges, Cys3-Cys24, CyS9-Cys29, Cys13-Cys31 and Cys19-Cys34, are present in the molecule.